We had previously found that cells from 6 human rhabdomyosarcoma (RMS) cell lines when injected in the subcutis of nude mice retain their initial cytology (embryonal vs. alveolar). Moreover, we found correlation between the histologic subtype (alveolar vs. embryonal) of RMS and the time/size of tumor growth in the subcutis of nude mice. In addition, all cell lines were found to express high levels of the c-myc oncogene, with or without c-myc amplification. N-myc was not expressed or amplified in any of the lines. In the present study, using the same RMS cell lines as in the previous study, we evaluated levels of expression of the c-myc oncogene in relation to (1) the histologic subtype, (2) tumor cell growth in vitro, and in the subcutis of nude mice, and (3) the ability of the tumor cells to form pulmonary nodules after tail vein injection mice. c-myc expression was evaluated by Northern blot analysis, and densitometric analysis was performed with an ultrascan XL laser densitometer. To determine if myc expression is independent of cell proliferation, we performed Northern blot analysis of the same cells after serum deprivation. In vitro tumor growth was evaluated by counting cells in a Coulti counter every 3 days for a total period of 12 days. In order to correlate c-myc expression with tumor growth, the results of tumor growth in the subcutis of nude mice from the previous study were used. The potential of tumor cells to cause pulmonary nodules ("metastases") is evaluated with tail vein injections of 2.5 x 105 cells from each RMS cell line into six mice per line. All mice were sacrificed after six weeks and autopsied. Pulmonary nodules were counted macroscopically and confirmed histologically.